1. Field of Invention
This invention relates to host cell proteins involved in retrovirus replication. More specifically, the invention relates to host cell proteins involved in human immunodeficiency virus (HIV-1) replication.
2. Description of Related Art
Retroviral agents have been implicated in a number of diseases, including cancer, autoimmune disease and AIDS. Human immunodeficiency virus (HIV) infection causes chronic progressive depletion of CD4+ T lymphocytes (CD4+ cells) and infection of macrophages, resulting in acquired immune deficiency syndrome.
Human immunodeficiency virus type 1 (HIV-1), a retrovirus, is the etiologic cause of AIDS. The HIV-1 envelope glycoprotein, gp120, specifically binds to the CD4 receptor on T lymphocytes and on monocytes and macrophages. Although infection of T lymphocytes requires cellular proliferation and DNA synthesis, productive infection of monocytes can occur independently of cellular DNA synthesis (Weinberg, J. B., et al, (1991) J. Exp. Med. 174:1477-82). When HIV-1 infects activated CD4+ lymphocytes, it is lethal, but infected monocytes are relatively resistant to destruction by the virus. Consequently, these cells, once infected with HIV-1, serve as long-lived reservoirs of the virus. Not only are these cells a source of replicating virus, but their virally-mediated dysfunction may contribute to increased susceptibility to opportunistic infections that are the hallmark of AIDS.
One of the current directions in anti-HIV-1 drug development focuses on the invariant host cell proteins involved in HIV-1 replication. Recently, a number of host cell proteins have been found to be necessary for HIV-1 replication and infectivity. In contrast, little is known about the proteins participating in the cellular defense against HIV-1 infection.
Feshchenko et al. identified TULA, a lymphoid SH3- and UBA domain-containing protein, which binds to ubiquitin and the ubiquitin-protein ligase c-Cbl (TULA: on SH3- and UBA-containing protein that binds to c-Cbl and ubiquitin. Oncogene 23, 4690-4706 (2004)). It was shown that TULA counteracts c-Cbl-dependent downregulation of protein tyrosine kinases (PTKs) and PTK-mediated signaling in a UBA-dependent fashion.
The gene encoding for TULA, which was termed UBASH3A, was examined by Wattenhofer et al. for possible involvement in autosomal recessive deafness; the authors concluded that UBASH3A plays no role in this disease [1]. The protein product of UBASH3A was not isolated or analyzed in this study, but the prediction based on the gene sequence showed that this protein possesses an SH3 and a UBA domain and a region of homology with some human, worm and fly proteins referred to as HCD (FIG. 1).
Feshchenko et al. identified the protein product of UBASH3A in the course of co-purification of c-Cbl with c-Cbl-associated proteins from the JMC-D3 clone of the human T-cell line Jurkat [2]. The ability of this protein to bind to c-Cbl via the interactions of its SH3 domain and the proline-rich region of c-Cbl was confirmed using co-immunoprecipitation and in vitro “pull-down” assays. Using Northern blotting, RT-PCR and Western blotting, expression of TULA in various cell lines and tissues were analyzed. It was demonstrated that TULA is preferentially expressed in lymphoid cells. Feshchenko et al. also showed that this protein binds well to ubiquitin via its UBA domain. Considering this finding and the predominant expression of this protein in T lymphocytes and other cells of lymphoid lineage, this protein was termed T-cell ubiquitin ligand (TULA). It was further demonstrated that TULA inhibits c-Cbl-dependent downregulation of protein tyrosine kinases (PTKs) [2].
Two other groups have published results related to TULA. Carpino et al. identified TULA, which they termed Sts-2, independently [3]. The results obtained by this group are similar to those described in [2] with regard to TULA expression, but are different with regard to TULA's interactions with other proteins. Carpino et al. failed to show TULA's binding to c-Cbl or stabilization of PTKs. A more recent study of TULA (Sts-2) by Kowanetz et al. [4] has corroborated the data described in [2] on c-Cbl binding and PTK stabilization.
The second protein of the TULA family (NP—116262, BAC11468), referred to as TULA-2 herein, is expressed ubiquitously [3-5]. This protein has been identified by Carpino et al. [5] and characterized by Carpino et al. [3] and Kowanetz et al. [4]. TULA-2 is similar to TULA in its ability to bind to c-Cbl and stabilize PTKs [4].
The above discussed studies do not describe the effect of TULA on HIV-1 biogenesis.
Studies of negative regulators of HIV-1 biogenesis and infectivity and were reported by Shindo et al., 2003 [6]; Mangeat et al., 2003 [7]; Zhang et al., 2003 [29]; Yap et al. [9], 2004; Carlson et al., 2004 [11]. However, none of these references characterize TULA proteins as regulators of HIV-1 biogenesis.
Several proteins have been shown to function as parts of the intracellular defense against HIV-1 infection, of which APOBEC proteins provide the best-studied example [6, 7]. APOBEC proteins are cytidine deaminases that can hypermutate nascent reverse transcripts, thus inhibiting the replication of HIV-1. However, recent findings suggest that the anti-HIV-1 effect of APOBEC proteins correlates with their ability to prevent the accumulation of reverse transcripts and not with the induction of hypermutation [8]. TRIM5α is another intracellular anti-HIV factor, which acts by promoting rapid, premature disassembly of retroviral capsids [9, 10]. OKT18, another antiviral factor, exerts its anti-HIV activity through suppression of HIV-1 transcription [11]. None of these factors can effectively suppress HIV-1 infection, since HIV-1 evades and actively counters their effects. For example, the effect of APOBEC is countered by the viral protein Vif [6, 7].
Based on the existing knowledge about cellular factors restricting HIV-1 infection, neither of these factors is related to TULA and none of their effects recapitulates the anti-HIV effect of TULA. Therefore, TULA represents a novel anti-HIV factor as discovered by the inventors.
Thus, despite the foregoing developments, there is a need in the art for cellular proteins working as natural barriers to retroviral infection and specifically to HIV-1 infection.
All references cited herein are incorporated herein by reference in their entireties.